Pancreatic ductal adenocarcinoma (PDAC) is an extremely lethal cancer with limited treatment options. This disease is characterized by a high frequency of activating KRAS mutations (95%), which is a known driver of PDAC progression. However, to date, no successful anti-K-Ras therapies have been developed. Current efforts have focused on inhibition of effectors of K-Ras signaling, in particular the Raf and PI3K signaling pathways. However, inhibitors targeting these pathways, when used as monotherapy or in combination, have been ineffectual for long-term treatment of KRAS mutant cancers. The lack of success of these inhibitors is due, in part, to the importance of other effectors in K-Ras-dependent cancer growth and the upregulation of compensatory signaling programs that overcome inhibitor activity. Consequently, there is a pressing need to better understand the role of other effector signaling events that support mutant K-Ras-driven PDAC growth in order to design effective combinatorial-targeted therapies. The small GTPase Rac1 has a known role in driving K-Ras mutant cancers, but the specific effectors through which Rac1 promotes tumor growth have not been defined. For my studies, we hypothesize that the PAK1 serine/threonine kinase, and related isoforms, PAK2 and PAK3, are key components downstream of Rac1 in mutant K-Ras PDAC. In support of this, my preliminary results found that Pak1 protein levels are overexpressed in pancreatic cancer cell lines and in patient tumor samples when compared to normal tissues. Additionally I determined that stable shRNA-mediated suppression of Pak1 protein expression inhibited PDAC anchorage-independent and -dependent growth and Matrigel invasion in vitro. Further, knockdown of K-Ras in PDAC cell lines results in reduced phospho-PAK1 (T423) levels, indicating a decrease in PAK1 activity. Recently, genetic suppression of PAK3 was determined to sensitive cells to pharmacologic inhibition of ERK1/2. This study suggests that other Group I PAK isoforms, PAK2 and PAK3, may be contributing to PAK1 driving PDAC growth. These results provide the rationale and foundation for my proposed studies to further validate the role of Group I PAK isoforms in PDAC. Additionally, because I have observed a nuclear sequestration of the normally cytoplasmic Pak1 in PDAC cells and patient tumor samples, I hypothesize that a Rac1-Pak1 activity is critical for supporting mutant K-Ras pancreatic cancer growth and invasion through distinct, subcellular localization-specific functions. To test this hypothesis, I will apply a recently designed inducible Pak1 construct for the spatio-temporal determination of Pak1 substrates that are important for Pak1-dependent PDAC growth. These studies will generate novel basic and translational information regarding PAK function and will require my application of a spectrum of biochemical, molecular and cellular techniques.